Clostridium septicum alpha toxin (CSA) plays significant roles in ruminant's braxy. Genetically engineered CSA has been shown to function as a potential vaccine candidate in the prevention of the disease caused by Clostridium septicum. In the present study, we synthesized a non-toxic recombinant, rCSA by introducing four amino acid substitutions (C86L/N296A/H301A/W342A) and 11-amino-acid deletion (residues 212 to 222). Compared to recombinant CSA, rCSA showed no cytotoxicity to MDCK cells and was not fatal to mice. Moreover, rCSA could protect immunized mice against 5?×?mouse LD (100% lethal dose) of crude CSA without obvious pathological change. Most importantly, rabbits immunized with rCSA produced high titers of neutralizing antibodies which protected the rabbits against crude CSA challenge. These data suggest that genetically detoxified rCSA is a potential subunit vaccine candidate against braxy.